The etiological agent of AIDS and ARC has variously been termed LAV, HTLV-III, ARV, and HIV. Hereinafter it will be referred to as HIV. Detection of the RNA or DNA of this virus is possible through a variety of probe sequences and hybridization formats.
PCT WO 88/01302, filed Aug. 11, 1987, discloses thirteen HIV oligonucleotides for use as probes in detecting HIV DNA or RNA. PCT WO 87/07906, filed Jun. 22, 1987, discloses variants of HIV viruses and the use of their DNA to diagnoses AIDS. EP 0 326 395 A2, filed Jan. 27, 1989, discloses an HIV DNA probe spanning nucleotides 2438-2457 for detecting sequences associated with multiple sclerosis.
The advent of the polymerase chain reaction has stimulated a range of assays using probes mainly from regions of the pol and gag genes. Spector et al. (Clin. Chem. 35/8:1581-1587, 1989) and Kellog et al. (Analytical Biochem 189:202-208, 1990) disclose a quantitative assay for HIV proviral DNA using polymerase chain reaction using a primer from the HIV gag gene. Lomell et al. (Clin. Chem. 35/9:1826-1831) disclose an amplifiable RNA probe complementary to a conserved region of the HIV pol gene mRNA. Coutlee et al. (Anal. Biochem. 181:96-105, 1989) disclose immunodetection of HIV DNA using the polymerase chain reaction with a set of primers complementary to sequences from the HIV pol and gag genes. EP 0 272 098, filed Dec. 15, 1987, discloses PCR amplification and detection of HIV RNA sequences using oligonucleotide probes spanning nucleotides 8538-8547 and 8658-8677. EP 0 229 701, filed Jan. 9, 1987 discloses detection of HIV by amplification of DNA from the HIV gag region. PCT WO 89/10979 discloses a nucleic acid probe assay combining amplification and solution hybridization using capture and reporter probes followed by immobilization on a solid support. A region within the gag p 17 region of HIV was amplified with this technique.
An alternative strategy is termed "reversible target capture." For example, Thompson et al. (Clin. Chem. 35/9:178-1881, 1989) disclose "reversible target capture" of HIV RNA, wherein a commercially available dA-tailed synthetic oligonucleotide provided selective purification of the analyte nucleic acid, and a labeled antisense RNA probe complementary to the HIV pol gene provided signal. Gillespie et al. (Molecular and Cellular Probes 3:73-86, 1989) discloses probes for reversible target capture of HIV RNA, wherein the probes are complementary to nucleotides 2094-4682 of the HIV pol gene.
Kumar et al. disclose a "probe shift" assay for HIV DNA, using DNA sequences complementary to the HIV gag and pol genes. The probe shift assay depends on the hybridization of a labeled oligonucleotide to a PCR-amplified segment in solution. The hemiduplex thereformed is detected following fractionation on nondenaturing gels.
Keller et al. (Anal. Biochem. 177:27-32, 1989) disclose a microtiter-based sandwich assay to detect HIV DNA spanning the Pst I site of the gag coding region.
Viscidi et al. (J. Clin. Micro. 27:120-125, 1989) disclose a hybridization assay for HIV RNA using a solid phase anti-biotin antibody and an enzyme-labeled monoclonal antibody specific for DNA-RNA hybrids, wherein the probe spanned nearly all of the polymerase gene and the 3' end of the gag gene.
European Patent Application (EPA) 89311862, filed Nov. 16, 1989 discloses a diagnostic kit and method using a solid capture means for detecting nucleic acid, and describes the use of DNA sequences complementary to the HIV gag gene to detect HIV DNA.
Commonly owned U.S. Pat. No. 4,868,105 issued Sep. 19, 1989, describes a solution phase nucleic acid sandwich hybridization assay in which analyte nucleic acid is first hybridized in solution to a labeling probe set and to a capturing probe set in a first vessel. The probe-analyte complex is then transferred to a second vessel that contains a solid-phase-immobilized probe that is complementary to a segment of the capturing probes. The segments hybridize to the immobilized probe, thus removing the complex from solution. Having the analyte in the form of an immobilized complex facilitates subsequent separation steps in the assay. Ultimately, single stranded segments of the labeling probe set are hybridized to labeled probes, thus permitting the analyte-containing complex to be detected via a signal generated directly or indirectly from the label.
Commonly owned European Patent Application (EPA) 883096976 discloses a variation in the assay described in U.S. Pat. No. 4,868,105 in which the signal generated by the labeled probes is amplified. The amplification involves the use of nucleic acid multimers. These multimers are branched polynucleotides that are constructed to have a segment that hybridizes specifically to the analyte nucleic acid or to a nucleic acid (branched or linear) that is bound to the analyte and iterations of a second segment that hybridize specifically to the labeled probe. In the assay employing the multimer, the initial steps of hybridizing the analyte to label or amplifier probe sets and capturing probe sets in a first vessel and transferring the complex to another vessel containing immobilized nucleic acid that will hybridize to a segment of the capturing probes are followed. The multimer is then hybridized to the immobilized complex and the labeled probes in turn hybridized to the second segment iterations on the multimer. Since the multimers provide a large number of sites for label probe attachment, the signal is amplified. Amplifier and capture probe sequences are disclosed for Hepatitis B virus, Neisseria gonorrhoeae, penicillin and tetracycline resistance in N. gonorrhoeae, and Chlamydia trachomatis.
Commonly owned copending application Ser. No. 558,897, filed Jul. 27, 1990, describes the preparation of large comb-type branched polynucleotide multimers for use in the above-described solution phase assay. The combs provide greater signal enhancement in the assays than the smaller multimers.
U.S. Pat. No. 5,030,557, issued Jul. 9, 1991, discloses a "helper" oligonucleotide selected to bind to the analyte nucleic acid and impose a different secondary and tertiary structure on the target to facilitate the binding of the probe to the target.